Histopathology is the diagnosis and study of diseases of the tissues and involves examining tissues and/or cells under a microscope. Histopathologists are responsible for making tissue diagnoses and helping clinicians manage a patient’s care.
What type of tissue is used by Histopathology.?
Four basic types of human tissue can be stained and viewed using various histological techniques. Epithelium, connective tissue, muscle tissue, and nervous tissue have commonalities but look very distinct structurally after staining.
Steps Of Histo-pathology
- Obtaining a fresh specimen. Fresh tissue specimens will come from various sources. …
- Fixation. The specimen is placed in a liquid fixing agent (fixative) such as formaldehyde solution (formalin).
- Dehydration.
- Clearing.
- Wax infiltration.
- Embedding or blocking out.
Step 1: Obtaining Fresh Specimen:
Fresh tissue specimens will come from various sources. It should be noted that they can very easily be damaged during removal from the patient or experimental animal. It is important that they are handled carefully and appropriately fixed as soon as possible after dissection.
Step 2: Fixation:
The specimen is placed in a liquid fixing agent (fixative) such as formaldehyde solution (formalin). This will slowly penetrate the tissue causing chemical and physical changes that will harden and preserve the tissue and protect it against subsequent processing steps.2 Generally, this will mean that the specimen should fix for between 6 and 24 hours. Most laboratories will use a fixative step as the first station on their processor.
Step 3: Dehydration:
Because melted paraffin wax is hydrophobic (immiscible with water), most of the water in a specimen must be removed before it can be infiltrated with wax
A typical dehydration sequence for specimens not more than 4mm thick would be:
- 70% Alcohol 2o hour’s or One Day
- 85% Alcohol 4 Hours
- 95% ethanol 4 hours
- Absolute Alcohol (1) 4 Hours
- Absolute Alcohol (2) 2 Hours
At this point, all but a tiny residue of tightly bound (molecular) water should have been removed from the specimen. ★ Step 4. Clearing:
Unfortunately, although the tissue is now essentially waterfree, we still cannot infiltrate it with wax because wax and ethanol are largely immiscible
A typical clearing sequence for specimens not more than 4mm thick would be:
xylene + Alcohol (50+50) 2 hours xylene || (15 minutes)
Step 5. Wax infiltration:
The tissue can now be infiltrated with a suitable histological wax. Although many different reagents have been evaluated and used for this purpose over many years, the paraffin wax-based histological waxes are the most popular. A typical wax is liquid at 60°C and can be infiltrated into tissue at this temperature then allowed to cool to 20°C, where it solidifies to a consistency that allows sections to be consistently cut.
A typical infiltration sequence for specimens not more than 4mm thick would be:
Paraffin wax (1) 15 min
Paraffin wax (2) 2 hours
6. Embedding or blocking out:
Now that the specimen is thoroughly infiltrated with wax, it must be formed into a “block” which can be clamped into a microtome for section cutting. This step is carried out using an “embedding center” where a mold is filled with molten wax and the specimen placed into it. The specimen is very carefully orientated in the mold because its placement will determine the “plane of section”, an important consideration in both diagnostic and research histology.
Apparatus Use in Histo-pathology
Why we do Histopathology:
Histopathology enables professionals to look for changes in cells that explain the actual cause of the patient’s illness. Pathologists are able to reach a diagnosis by examining a small piece of tissue from various organs.
Histopathology is vital as it broadens and progresses treatments options.
I am Zain Ul Abedin.I am working for the welfare of the Poultry industry. I wrote many articles on different Poultry diseases in many National and local newspapers and also in veterinary News. I am a Member Pakistan vet care Association.
